Transdermal Delivery of Kidney-Targeting Nanoparticles Using Dissolvable Microneedles

Abstract

Introduction: Chronic kidney disease (CKD) affects approximately 13% of the world's population and will lead to dialysis or kidney transplantation. Unfortunately, clinically available drugs for CKD show limited efficacy and toxic extrarenal side effects. Hence, there is a need to develop targeted delivery systems with enhanced kidney specificity that can also be combined with a patient-compliant administration route for such patients that need extended treatment. Towards this goal, kidney-targeted nanoparticles administered through transdermal microneedles (KNP/MN) is explored in this study.

Methods: A KNP/MN patch was developed by incorporating folate-conjugated micelle nanoparticles into polyvinyl alcohol MN patches. Rhodamine B (RhB) was encapsulated into KNP as a model drug and evaluated for biocompatibility and binding with human renal epithelial cells. For MN, skin penetration efficiency was assessed using a Parafilm model, and penetration was imaged via scanning electron microscopy. In vivo, KNP/MN patches were applied on the backs of C57BL/6 wild type mice and biodistribution, organ morphology, and kidney function assessed.

Results: KNP showed high biocompatibility and folate-dependent binding in vitro, validating KNP's targeting to folate receptors in vitro. Upon transdermal administration in vivo, KNP/MN patches dissolved within 30 min. At varying time points up to 48 h post-KNP/MN administration, higher accumulation of KNP was found in kidneys compared with MN that consisted of the non-targeting, control-NP. Histological evaluation demonstrated no signs of tissue damage, and kidney function markers, serum blood urea nitrogen and urine creatinine, were found to be within normal ranges, indicating preservation of kidney health.

Conclusions: Our studies show potential of KNP/MN patches as a non-invasive, self-administrable platform to direct therapies to the kidneys.

Keywords: Chronic kidney disease; Folate; Kidney-targeting; Microneedles; Nanoparticles.

Figure 1

(a) Fabrication of dissolvable MN patch using micromolding technique. (b) Photograph of dissolvable MN patch. (c) SEM images of the MN array (inset: single MN). The MN patch consist of a 21 × 21 needle array, and each MN has a 300 μm base width to prevent significant tissue damage and 600 μm length for penetration into the dermis. (d) Dissolution efficiency and RhB release profile from the MN patch in PBS (10 mL, pH 7.4) for up to 10 min at 37 °C. Each data point represents the mean RhB fluorescence intensity at 556 nm excitation and 580 nm emission spectrum peak wavelength. Inset of d shows the light microscopic images of MN patch captured at different time points.

Figure 2

(a) The insertion efficiency generated in each Parafilm M® layer upon MN patch insertion. SEM images of MN insertion efficiency in a Parafilm M® model with varying layers: (b) 2 layers, (c) 4 layers, (d) 6 layers, and (e) 8 layers.

Figure 3

(a) Schematic representation of micelle self-assembly and micelle loading into MN. Portions were adapted and reprinted from Servier Medical Art, under creative commons license. TEM micrographs of (b) control (non-targeting) NP and KNP with (c) low folate content (20FA-KNP) and (d) high folate content (40FA-KNP).

Figure 4

DLS measurements of particle size distributions of (a) non-targeting NP, (b) 20FA-KNP and (c) 40FA-KNP in the MN polymer solution shows recovery of KNP (n = 4).

Figure 5

In vitro cell viability studied by MTT assay of (a) RPTEC and (b) WT 9–12. Cells were treated with 1–100 µM of non-targeting NP, 20FA-KNP and 40FA-KNP for 24 h. % viability normalized by PBS-treated controls. Cellular binding of RhB-loaded non-targeting NP, 20FA-KNP, and 40FA-KNP in (c) RPTEC and (d) WT 9–12. Cells were treated with 50 µM NP for 1 and 2 h with or without excess free (5 mM) folate (n = 6).

Figure 6

Biodistribution study of KNP, NP, and PBS after 24 and 48 h in C57BL/6 mice. (a) Ex vivo organ imaging post-MN application shows accumulation of KNP mostly in the kidney, liver, intestines, and skin. (b) Quantitative measurements of fluorescence levels in the brain, heart, lung, kidney, liver, spleen, bladder, intestines, and skin post-MN application at 24 and 48 h and PBS-treated mice via IP injection at 48 h. (c) Quantitative comparison of kidney fluorescence levels showed an average of 2.2 times higher accumulation of KNP compared to non-targeting NP after 24 h, and an average of 7.7 times higher accumulation of KNP compared to non-targeting NP after 48 h. (d) The half-life of RhB was found to be 90 ± 12 min for KNP/MN, and 120 ± 10 min for NP/MN (n = 4).

Figure 7

H & E staining of organs after 24 and 48 h after KNP/MN, NP/MN, and PBS administration displayed no morphological changes. Scale bar: 100 µm.